Dimethyl-Dioctadecyl-Ammonium Bromide/Poly(lactic acid) Nanoadjuvant Enhances the Immunity and Cross-Protection of an NM2e-Based Universal Influenza Vaccine.

For most frequent respiratory viruses, there is an urgent need for a universal influenza vaccine to provide cross-protection against intra- and heterosubtypes. We previously developed an Escherichia coli fusion protein expressed extracellular domain of matrix 2 (M2e) and nucleoprotein, named NM2e, and then combined it with an aluminum adjuvant, forming a universal vaccine. Although NM2e has demonstrated a protective effect against the influenza virus in mice to some extent, further improvement is still needed for the induction of immune responses ensuring adequate cross-protection against influenza. Herein, we fabricated a cationic solid lipid nanoadjuvant using poly(lactic acid) (PLA) and dimethyl-dioctadecyl-ammonium bromide (DDAB) and loaded NM2e to generate an NM2e@DDAB/PLA nanovaccine (Nv). In vitro experiments suggested that bone marrow-derived dendritic cells incubated with Nv exhibited ∼4-fold higher antigen (Ag) uptake than NM2e at 16 h along with efficient activation by NM2e@DDAB/PLA Nv. In vivo experiments revealed that Ag of the Nv group stayed in lymph nodes (LNs) for more than 14 days after initial immunization and DCs in LNs were evidently activated and matured. Furthermore, the Nv primed T and B cells for robust humoral and cellular immune responses after immunization. It also induced a ratio of IgG2a/IgG1 higher than that of NM2e to a considerable extent. Moreover, NM2e@DDAB/PLA Nv quickly restored body weight and improved survival of homo- and heterosubtype influenza challenged mice, and the cross-protection efficiency was over 90%. Collectively, our study demonstrated that NM2e@DDAB/PLA Nv could offer notable protection against homo- and heterosubtype influenza virus challenges, offering the potential for the development of a universal influenza vaccine.

Pitch Improvement in Attentional Blink: A Study across Audiovisual Asymmetries.

Attentional blink (AB) is a phenomenon in which the perception of a second target is impaired when it appears within 200-500 ms after the first target. Sound affects an AB and is accompanied by the appearance of an asymmetry during audiovisual integration, but it is not known whether this is related to the tonal representation of sound. The aim of the present study was to investigate the effect of audiovisual asymmetry on attentional blink and whether the presentation of pitch improves the ability to detect a target during an AB that is accompanied by audiovisual asymmetry. The results showed that as the lag increased, the subject's target recognition improved and the pitch produced further improvements. These improvements exhibited a significant asymmetry across the audiovisual channel. Our findings could contribute to better utilizations of audiovisual integration resources to improve attentional transients and auditory recognition decline, which could be useful in areas such as driving and education.

Rational design of a 'two-in-one' immunogen DAM drives potent immune response against mpox virus.

The global outbreak of the mpox virus (MPXV) in 2022 highlights the urgent need for safer and more accessible new-generation vaccines. Here, we used a structure-guided multi-antigen fusion strategy to design a 'two-in-one' immunogen based on the single-chain dimeric MPXV extracellular enveloped virus antigen A35 bivalently fused with the intracellular mature virus antigen M1, called DAM. DAM preserved the natural epitope configuration of both components and showed stronger A35-specific and M1-specific antibody responses and in vivo protective efficacy against vaccinia virus (VACV) compared to co-immunization strategies. The MPXV-specific neutralizing antibodies elicited by DAM were 28 times higher than those induced by live VACV vaccine. Aluminum-adjuvanted DAM vaccines protected mice from a lethal VACV challenge with a safety profile, and pilot-scale production confirmed the high yield and purity of DAM. Thus, our study provides innovative insights and an immunogen candidate for the development of alternative vaccines against MPXV and other orthopoxviruses.

Clinical and molecular epidemiology of enterovirus D68 from 2013 to 2020 in Shanghai.

Enterovirus D68 (EV-D68) is an emerging pathogen that has caused outbreaks of severe respiratory disease worldwide, especially in children. We aim to investigate the prevalence and genetic characteristics of EV-D68 in children from Shanghai. Nasopharyngeal swab or bronchoalveolar lavage fluid samples collected from children hospitalized with community-acquired pneumonia were screened for EV-D68. Nine of 3997 samples were EV-D68-positive. Seven of nine positive samples were sequenced and submitted to GenBank. Based on partial polyprotein gene (3D) or complete sequence analysis, we found the seven strains belong to different clades and subclades, including three D1 (detected in 2013 and 2014), one D2 (2013), one D3 (2019), and two B3 (2014 and 2018). Overall, we show different clades and subclades of EV-D68 spread with low positive rates (0.2%) among children in Shanghai between 2013 and 2020. Amino acid mutations were found in the epitopes of the VP1 BC and DE loops and C-terminus; similarity analysis provided evidence for recombination as an important mechanism of genomic diversification. Both single nucleotide mutations and recombination play a role in evolution of EV-D68. Genetic instability within these clinical strains may indicate large outbreaks could occur following cumulative mutations.

Rapid identification of full-length genome and tracing variations of monkeypox virus in clinical specimens based on mNGS and amplicon sequencing.

The monkeypox virus (MPXV) has triggered a current outbreak globally. Genome sequencing of MPXV and rapid tracing of genetic variants will benefit disease diagnosis and control. It is a significant challenge but necessary to optimize the strategy and application of rapid full-length genome identification and to track variations of MPXV in clinical specimens with low viral loads, as it is one of the DNA viruses with the largest genome and the most AT-biased, and has a significant number of tandem repeats. Here we evaluated the performance of metagenomic and amplicon sequencing techniques, and three sequencing platforms in MPXV genome sequencing based on multiple clinical specimens of five mpox cases in Chinese mainland. We rapidly identified the full-length genome of MPXV with the assembly of accurate tandem repeats in multiple clinical specimens. Amplicon sequencing enables cost-effective and rapid sequencing of clinical specimens to obtain high-quality MPXV genomes. Third-generation sequencing facilitates the assembly of the terminal tandem repeat regions in the monkeypox virus genome and corrects a common misassembly in published sequences. Besides, several intra-host single nucleotide variations were identified in the first imported mpox case. This study offers an evaluation of various strategies aimed at identifying the complete genome of MPXV in clinical specimens. The findings of this study will significantly enhance the surveillance of MPXV.

Long-lasting humoral and cellular memory immunity to vaccinia virus Tiantan provides pre-existing immunity against mpox virus in Chinese population.

Investigating immune memory to vaccinia virus and pre-existing immunity to mpox virus (MPXV) among the population is crucial for the global response to this ongoing mpox epidemic. Blood was sampled from vaccinees inoculated with vaccinia virus Tiantan (VTT) strain born before 1981 and unvaccinated control subjects born since 1982. After at least 40 years of the inoculation, 60% or 5% VTT vaccinees possess neutralizing antibodies (NAbs) to VTT or MPXV, with at least 50% having T cell memory to VTT protein antigens. Notably, 46.7% vaccinees show pre-existing T cell responses to MPXV. Broad pre-existing CD8+ T cell reactivities to MPXV are detected not only against conserved epitopes but also against variant epitopes between VTT and MPXV. Persistent NAbs and T cell memory to VTT among vaccinees, along with pre-existing T cells to MPXV among both vaccinees and the unvaccinated population, indicate a particular immune barrier to mpox.

Mpox (formerly monkeypox): pathogenesis, prevention, and treatment.

In 2022, a global outbreak of Mpox (formerly monkeypox) occurred in various countries across Europe and America and rapidly spread to more than 100 countries and regions. The World Health Organization declared the outbreak to be a public health emergency of international concern due to the rapid spread of the Mpox virus. Consequently, nations intensified their efforts to explore treatment strategies aimed at combating the infection and its dissemination. Nevertheless, the available therapeutic options for Mpox virus infection remain limited. So far, only a few numbers of antiviral compounds have been approved by regulatory authorities. Given the high mutability of the Mpox virus, certain mutant strains have shown resistance to existing pharmaceutical interventions. This highlights the urgent need to develop novel antiviral drugs that can combat both drug resistance and the potential threat of bioterrorism. Currently, there is a lack of comprehensive literature on the pathophysiology and treatment of Mpox. To address this issue, we conducted a review covering the physiological and pathological processes of Mpox infection, summarizing the latest progress of anti-Mpox drugs. Our analysis encompasses approved drugs currently employed in clinical settings, as well as newly identified small-molecule compounds and antibody drugs displaying potential antiviral efficacy against Mpox. Furthermore, we have gained valuable insights from the process of Mpox drug development, including strategies for repurposing drugs, the discovery of drug targets driven by artificial intelligence, and preclinical drug development. The purpose of this review is to provide readers with a comprehensive overview of the current knowledge on Mpox.

Immunogenicity and Safety of a SARS-CoV-2 Inactivated Vaccine KCONVAC in Chinese Children: Randomized, Double-blind, Placebo-controlled Phase 1 and 2 Trials.

BACKGROUND: It is important to extend the indication of severe acute respiratory syndrome coronavirus 2 vaccine to children to improve the vaccine intake rate and reduce infection in this population. METHODS: In 2 phase 1 and phase 2 randomized, double-blind and placebo-controlled trials, 84 and 480 Chinese healthy children 3 to 17 years old were enrolled, respectively, and randomized in 3:1 ratio to receive 2 doses of a severe acute respiratory syndrome coronavirus 2 inactivated vaccine, KCONVAC or placebo. The 2 doses were given 28 days apart. Adverse events (AEs) were recorded through Day 28 after each dosing. Live virus neutralizing antibody and receptor binding domain antibody (RBD-IgG) were tested before vaccination and after the second dose. RESULTS: Two doses of the vaccine, KCONVAC, elicited geometric mean titers of 142-150 for neutralizing antibody and 4154-4253 for RBD-IgG 28 days after the second dose. Seroconversion rates were 100% after 2 doses for both antibodies in both trials. The predominant AEs were injection-site pain, cough and fever. Most AEs were grade 1 or 2 in intensity. Five participants reported 6 vaccination-unrelated serious AEs in the phase 2 trial. CONCLUSIONS: Two doses of this study vaccine, KCONVAC, were well tolerated and immunogenic in children 3 to 17 years of age.

Molecular Dynamics Simulation and Viscosity Analysis of Red Mud-Steel Slag Glass-Ceramics.

The preparation of glass-ceramics with red mud and steel slag can not only solve the pollution problem caused by industrial waste slag but also produce economic benefits. It is difficult to analyze the high-temperature melt with the existing test methods, so the simulation experiment with molecular dynamics calculation becomes an important research method. The effects of steel slag content on the microstructure of red mud glass-ceramics were studied by molecular dynamics method. The results show that the binding ability of Si-O, Al-O, and Fe-O decreases with the increase in steel slag content. The number of Si-O-Si bridge oxygen increased gradually, while the number of Al-O-Al, Al-O-Fe, and Fe-O-Fe bridge oxygen decreased significantly. The number of tetrahedrons [SiO4] increased, the number of tetrahedrons [FeO4] and [AlO4] decreased, and the total number of three tetrahedrons decreased. The mean square displacement value of Si4+ and O2- increases first and then decreases, resulting in the viscosity of the system decreasing first and then increasing. The molecular dynamics method is used to analyze the structure of red mud-steel slag glass-ceramics on the microscopic scale, which can better understand the role of steel slag and has guiding significance for the experiment of this kind of glass-ceramics.

Rapid and sensitive one-tube detection of mpox virus using RPA-coupled CRISPR-Cas12 assay.

Mpox is caused by a zoonotic virus belonging to the Orthopoxvirus genus and the Poxviridae family. In this study, we develop a recombinase polymerase amplification (RPA)-coupled CRISPR-Cas12a detection assay for the mpox virus. We design and test a series of CRISPR-derived RNAs(crRNAs) targeting the conserved D6R and E9L genes for orthopoxvirus and the unique N3R and N4R genes for mpox viruses. D6R crRNA-1 exhibits the most robust activity in detecting orthopoxviruses, and N4R crRNA-2 is able to distinguish the mpox virus from other orthopoxviruses. The Cas12a/crRNA assay alone presents a detection limit of 108 copies of viral DNA, whereas coupling RPA increases the detection limit to 1-10 copies. The one-tube RPA-Cas12a assay can, therefore, detect viral DNA as low as 1 copy within 30 min and holds the promise of providing point-of-care detection for mpox viral infection.

Discovery and identification of a novel canine coronavirus causing a diarrhea outbreak in Vulpes.

Cross-species transmission of viruses from wildlife animal reservoirs, such as bats, poses a threat to human and domestic animal health. Previous studies have shown that domestic animals have important roles as intermediate hosts, enabling the transmission of genetically diverse coronaviruses from natural hosts to humans. Here, we report the identification and characterization of a novel canine coronavirus (VuCCoV), which caused an epidemic of acute diarrhea in Vulpes (foxes) in Shenyang, China. The epidemic started on November 8, 2019, and caused more than 39,600 deaths by January 1, 2022. Full-length viral genomic sequences were obtained from 15 foxes with diarrhea at the early stage of this outbreak. The VuCCoV genome shared more than 90% nucleotide identity with canine coronavirus (CCoV) for three of the four structural genes, with the S gene showing a larger amount of divergence. In addition, 67% (10/15) of the VuCCoV genomes contained an open reading frame (ORF3) gene, which was previously only detected in CCoV-I genomes. Notably, VuCCoV had only two to three amino acid differences at the partial RNA-dependent RNA polymerase (RdRp) level to bat CoV, suggesting a close genetic relationship. Therefore, these novel VuCCoV genomes represent a previously unsampled lineage of CCoVs. We also show that the VuCCoV spike protein binds to canine and fox aminopeptidase N (APN), which may allow this protein to serve as an entry receptor. In addition, cell lines were identified that are sensitive to VuCCoV using a pseudovirus system. These data highlight the importance of identifying the diversity and distribution of coronaviruses in domestic animals, which could mitigate future outbreaks that could threaten livestock, public health, and economic growth.

Co(II)-Based Metal-Organic Framework Derived CA-CoNiMn-CLDHs with Peroxidase-like Activity for Colorimetric Detection of Phenol.

Given the serious harm of toxic phenol to human health and the ecological environment, it is urgent to develop an efficient, low-cost and sensitive nanoenzyme-based method to monitor phenol. MOF-derived nanozyme has attracted wide interest due to its hollow polyhedra structure and porous micro-nano frameworks. However, it is still a great challenge to synthesize MOF-derived multimetal synergistic catalytic nanoenzymes in large quantities with low cost. Herein, we reported the synthetic strategy of porous hollow CA-CoNiMn-CLDHs with ZIF-67 as templates through a facile solvothermal reaction. The prepared trimetallic catalyst exhibits excellent peroxidase-like activity to trigger the oxidative coupling reaction of 4-AAP and phenol in the presence of H2O2. The visual detection platform for phenol based on CA-CoNiMn-CLDHs is constructed, and satisfactory results are obtained. The Km value for CA-CoNiMn-CLDHs (0.21 mM) is lower than that of HRP (0.43 mM) with TMB as the chromogenic substrate. Because of the synergistic effect of peroxidase-like activity and citric acid functionalization, the built colorimetric sensor displayed a good linear response to phenol from 1 to 100 µM with a detection limit of 0.163 µM (3σ/slope). Additionally, the CA-CoNiMn-CLDHs-based visual detection platform possesses high-chemical stability and excellent reusability, which can greatly improve economic benefits in practical applications.

Single-Dose Intranasal Immunisation with Novel Chimeric H1N1 Expressing the Receptor-Binding Domain of SARS-CoV-2 Induces Robust Mucosal Immunity, Tissue-Resident Memory T Cells, and Heterologous Protection in Mice.

Current COVID-19 vaccines can effectively reduce disease severity and hospitalisation; however, they are not considerably effective in preventing infection and transmission. In this context, mucosal vaccines are pertinent to prevent SARS-CoV-2 infection and spread. In this study, we generated a replication-competent recombinant chimeric influenza A virus (IAV) expressing the receptor-binding domain (RBD) of a SARS-CoV-2 prototype in the C-terminus of the neuraminidase (NA) of A/Puerto Rico/08/1934 H1N1 (PR8). The remaining seven segments from A/WSN/1933 H1N1 (WSN) were named PR8NARBD/WSN. We observed that the recombinant virus with the WSN backbone demonstrated improved expression of NA and RBD. A single intranasal dose of PR8NARBD/WSN(103PFU) in mice generated robust mucosal immunity, neutralising antibodies, cellular immunity, and tissue-resident memory T cells specific to SARS-CoV-2 and IAV. Importantly, immunisation with PR8NARBD/WSN viruses effectively protected mice against lethal challenges with H1N1, H3N2 IAV, and SARS-CoV-2 Beta variant and significantly reduced lung viral loads. Overall, our research demonstrates the promising potential of PR8NARBD/WSN as an attractive vaccine against emerging SARS-CoV-2 variants and influenza A virus infections.

An IgM-like inhalable ACE2 fusion protein broadly neutralizes SARS-CoV-2 variants.

Many of the currently available COVID-19 vaccines and therapeutics are not effective against newly emerged SARS-CoV-2 variants. Here, we developed the metallo-enzyme domain of angiotensin converting enzyme 2 (ACE2)-the cellular receptor of SARS-CoV-2-into an IgM-like inhalable molecule (HH-120). HH-120 binds to the SARS-CoV-2 Spike (S) protein with high avidity and confers potent and broad-spectrum neutralization activity against all known SARS-CoV-2 variants of concern. HH-120 was developed as an inhaled formulation that achieves appropriate aerodynamic properties for rodent and monkey respiratory system delivery, and we found that early administration of HH-120 by aerosol inhalation significantly reduced viral loads and lung pathology scores in male golden Syrian hamsters infected by the SARS-CoV-2 ancestral strain (GDPCC-nCoV27) and the Delta variant. Our study presents a meaningful advancement in the inhalation delivery of large biologics like HH-120 (molecular weight (MW) ~ 1000 kDa) and demonstrates that HH-120 can serve as an efficacious, safe, and convenient agent against SARS-CoV-2 variants. Finally, given the known role of ACE2 in viral reception, it is conceivable that HH-120 has the potential to be efficacious against additional emergent coronaviruses.

The First Human Infection Case of Macacine Alphaherpesvirus 1 Virus in China.

BACKGROUND: Macacine alphaherpesvirus 1 (BV) was first reported in the 1930s and only about 60 cases have been diagnosed since then. METHODS: A 53-year-old male who worked as a veterinary surgeon, developed a fever with nausea and vomiting in April 2021 in Beijing, China. Real-time polymerase chain reaction (PCR) and metagenomics Next Generation Sequencing (mNGS) were used for diagnosis. RESULTS: BV DNA was confirmed by mNGS and PCR. The case died 51 days after onset, due to the damage to the brain and spinal cord caused by a viral infection and hypoxic-ischemic encephalopathy. The typical BV inclusion bodies in the brain were found for the first time. CONCLUSIONS: Here we reported the first human infection case of BV in China. This fatal case highlights the potential threat of BV to occupational workers and the essential role of surveillance.

Clinical, epidemiological, and virological features of Mpox in Beijing, China - May 31-June 21, 2023.

The first locally acquired case in the Chinese mainland was reported on May 31, 2023, lagging behind other countries. In this study, we aimed to examine the early clinical and epidemiological characteristics of the earliest cases of Mpox in Beijing, China. Additionally, we investigated the sequence and transmission patterns of the Mpox virus (MPXV). We analyzed 37 reported cases of Mpox in Beijing from May 31, 2023 to June 21, 2023. The age range of the subjects was 24-51 years. Thirty-six cases (97.3%) were identified in men who have sex with men (MSM), and 19 cases (51.4%) tested positive for the human immunodeficiency virus. Thirty-three cases were symptomatic, while four were asymptomatic. Skin lesions were observed in 32 cases (97.0%), fever in 26 (78.8%), and swollen lymph nodes in 17 (51.5%). Rash typically appeared in the genital or perianal area 1-3 days before fever onset, with a minimum incubation period of 2 days. For individuals with skin rashes, the skin lesion samples showed 100% positivity and low Ct values. There were high oropharyngeal swab (75.8%) and blood (84.6%) positivity rates. All MPXV strains belonged to the B.1.3 branch of the West African lineage. These strains carried 76-86 nucleotide substitutions compared with the reference human MPXV genome, and genetic diversity was observed. Our findings provide the first insights into the landscape of early transmission of Mpox in Beijing and help inform policy formulation in the Chinese mainland.

Neutralizing antibody levels associated with injectable and aerosolized Ad5-nCoV boosters and BA.2 infection.

BACKGROUND: Several COVID-19 vaccines are in widespread use in China. Few data exist on comparative immunogenicity of different COVID-19 vaccines given as booster doses. We aimed to assess neutralizing antibody levels raised by injectable and inhaled aerosolized recombinant adenovirus type 5 (Ad5)-vectored COVID-19 vaccine as a heterologous booster after an inactivated COVID-19 vaccine two-dose primary series. METHODS: Using an open-label prospective cohort design, we recruited 136 individuals who had received inactivated vaccine primary series followed by either injectable or inhaled Ad5-vectored vaccine and measured neutralizing antibody titers against ancestral SARS-CoV-2 virus and Omicron BA.1 and BA.5 variants. We also measured neutralizing antibody levels in convalescent sera from 39 patients who recovered from Omicron BA.2 infection. RESULTS: Six months after primary series vaccination, neutralizing immunity against ancestral SARS-CoV-2 was low and neutralizing immunity against Omicron (B.1.1.529) was lower. Boosting with Ad5-vectored vaccines induced a high immune response against ancestral SARS-CoV-2. Neutralizing responses against Omicron BA.5 were ≥ 80% lower than against ancestral SARS-CoV-2 in sera from prime-boost subjects and in convalescent sera from survivors of Omicron BA.2 infection. Inhaled aerosolized Ad5-vectored vaccine was associated with greater neutralizing titers than injectable Ad5-vectored vaccine against ancestral and Omicron SARS-CoV-2 variants. CONCLUSIONS: These findings support the current strategy of heterologous boosting with injectable or inhaled Ad5-vectored SARS-CoV-2 vaccination of individuals primed with inactivated COVID-19 vaccine.